Abstract Capsule is recognized as one of the most important structures of bacterial pathogens and has a long list of properties that contribute to successful pathogenesis within an infected host. Recent studies from our lab have revealed a novel role for the bacterial capsule locus in transmission between hosts. In addition, we have preliminary results demonstrating the capsule of Bordetella bronchiseptica has a profound effect on growth and persistence within the respiratory tract of its mammalian host. This project will explore the mechanistic details of these important effects by probing both the genetic and enzymatic basis for capsule assembly as well as its biochemical structure. We propose the following three aims: 1. Determine the role of each gene in the capsule (tEPS) locus in distinct stages of transmission. We will construct in-frame deletion of each of the 18 genes of the transmission associated extra-polysaccharide (tEPS) capsule locus to define the role of each gene in the profound effect on transmission that we observed by deleting this locus. We will use our innovative new assay to distinguish the genes that effect the shedding of bacteria from infected host from genes that effect the colonization of exposed animals. 2. Define the contributions of key genes to the structure and physiochemical properties of tEPS. We will isolate, purify and study the individual mutant EPS using a combined methodology (MS and NMR platforms) to determine the sugar sequences, linkage and structure. Relating the chemical structures and physiochemical properties of the EPS generated by wild type and various mutants that fail to transmit (in Aim 1) will reveal the specific aspects of the tEPS that are required for transmission. 3. Determine the enzymatic activities encoded by genes involved in transmission. Genes determined to be required for transmission in Aim 1 encode potential targets for interventions; blocking their enzymatic activity could prevent transmission. We will therefore express these genes in E. coli and define the enzymatic activities and kinetics of the critical steps. This project will reveal the chemical structure of a new capsule and identify key elements of the structure that contribute to the critical ability of pathogens to transmit between hosts, a key point of intervention to prevent the spread of infectious disease. It will also reveal the pathways involved in its assembly, thereby identifying targets for rational design of targeted treatments. Using a novel mouse model to study transmission and understanding how capsular polysaccharide(s) are being used by respiratory pathogens to transmit, are the noteworthy features of this proposal and highly relevant to groups investigating infectious bacterial diseases of the respiratory system.